See
“An Overview” for more information on the
derivation of these cell types.

Live
cells for distribution are sent frozen (in dry ice) in ampoules containing 5 x
10e5 or 1 x 10e6 cells. Under special circumstances, we can arrange to send
growing cells in flasks. For some cell types, RNA is also available upon
specific request.

See
"REVIEW
section VIII." for more information on cell shipments. Note: I’ve been developing this system for over 30 years; it’s complicated!
Since I don’t expect others to know which of the many cell types generated would be best
for their studies, I recommend that you consult with me first about what available cells
may be most appropriate.

We
have large batches of pre-stasis HMEC to distribute at passages 4-8; most
growth ceases around passages 13-16.Lower passage amps may be sent to allow growth of stocks in the
recipient’s lab. Pre-stasis HMEC contain a range of phenotypes, particularly at
lower passages; cells with luminal or progenitor markers make up ~5-30% of the
population.Pre-stasis HMEC are
genomically stable, even at stasis.I
most commonly distribute cells from specimens 184, 48R, and 240L (ages
16-21).More limited quantities of HMEC
derived from older woman are also available.

Very limited quantities of the BaP cultures are available. These include: 184Aa (p16 mutated), the precursor of 184A1 and other 184A- lines; 184Be (p16 silenced), the precursor of 184BE1 and 184BE2 lines, and 184Ce, the precursor of the184CeMY line. Talk with me directly about these cells. More 184Aa is available than any other BaP type. These cells were exposed to BaP and so harbor many point mutations.

We
have large batches of post-selection HMEC frozen at around passages 7-10 that
we distribute. Depending upon the individual, these cells cease active growth
around passages 14-25 (about 3 PD per passage).These cells are routinely available from women of different ages.I most commonly distribute cells from
specimens 184, 48R, 239, 240L and 161.

These cells are not normal.They have overcome the stasis barrier and show many differences from normal pre-stasis HMEC (e.g., in gene expression and promoter methylation), some of which are also seen in breast cancer cells. They express a generally basal phenotype, although some luminal marker may also be present at higher passages (e.g., mucins and keratin 18), and they exhibit metaplastic properties (Sauder et al. BMC Biology 2014). They become genomically unstable as they approach the telomere dysfunction senescence barrier. Post-selection HMEC have also been referred to as vHMEC, and suggested to give rise to metaplastic cancer (Keller et al. PNAS 2012). This is the cell type sold commercially as “normal primaries” (e.g., Lonza CC-2551 and Life Technologies A10565) although these significantly aberrant cells are neither normal nor primary.

In
2 separate experiments pre-stasis 184 HMEC were transduced with a genetic
suppressor element (GSE22) that produces a peptide that interferes with p53
function.In both cases there were a
few clonal outgrowths when the vast majority of the cells ceased growth at
stasis.These cells have not been
well-characterized.They express low but
detectable p16. These cells are not routinely distributed.

Pre-stasis 184, 240L, 122L, and 805P HMEC were transduced with shRNA to p16 at early passages (184F batch in M85 medium; the others in M87A+X medium). These cells have not been well-characterized. They express increased telomerase (TRAP) activity compared to the parental pre-stasis HMEC. These cells are not routinely distributed but can be made available.

We
have available for distribution stocks of fibroblast cells from several
reduction mammoplasty specimens for which HMEC are available, e.g., specimens
184, 48, 240, and 161. These cells are grown in a serum-containing medium.In theory, fibroblast stocks can be obtained
from any of our specimens, including the mastectomy derived tissues, but we
have not grown up stocks to distribute from more than a few.Frozen cells are available around passages
5-8, and they senesce around passages 12-20 (2-3 PD per passage) depending upon
the individual.These cells grow slower
than the HMEC, and we do not generally have as large stocks available.

We
have very limited quantities of
frozen organoids, and are therefore very reluctant to distribute any of this
material, but will make exceptions for specific studies. More primary tissue is
available from reduction mammoplasties than mastectomies. Talk with me directly
about these cells.

Unlimited
quantities of later passages of this immortal cell line are available for
distribution.Very limited quantities of
early passage newly immortal (pre-conversion) cells are available upon specific
request.Clonal isolates are also
available.These cells are wild type for
p53 and RB, and are not anchorage independent or tumorigenic.They have the most stable karyotype of our BaP-exposed immortally transformed lines. This line is derived from the BaP post-stasis precursor 184Aa (p16 mutated).

184A1was
transduced with GSE22.When the GSE22
is introduced pre-conversion (passage 12) conversion proceeds rapidly. When the
GSE22 is introduced into fully immortal 184A1 it can provide a matched p53(-)
culture to the p53(+) 184A1. These cells are not routinely distributed but can
be made available.

Early passage pre-conversion (passage 12) 184A1 was transduced with either the HPV16 -E6, -E7, SV40T, or E1A genes. These viral oncogenes produce numerous effects that may accelerate or alter the conversion process. These cells are not routinely distributed.

Unlimited
quantities of later passages of this immortal line are available for
distribution.Early passage cells are
not available. These cells are wild type for p53, and RB, and are not anchorage
independent or tumorigenic.They have
many karyotypic abnormalities. This line is derived from the BaP post-stasis precursor 184Aa.

Unlimited
quantities of this p53(-/-) immortal line are available for distribution. These
cells are wild type for RB, and have AIG.The karyotype is unstable.184AA2 is derived from the same BaP post-stasis precursor population (184Aa) as 184A1 and 184AA4. 184Aa was transduced with retroviral vectors that gave insertional mutagenesis at the p53 locus. We presume this line acquired addition errors at crisis.

Unlimited quantities of this p53(-/-) immortal line are available for distribution. These cells are wild type for RB, and have AIG by passage 50. The karyotype is unstable. 184AA3 is derived from the same BaP post-stasis precursor population (184Aa) as 184A1 and 184AA4. 184Aa was transduced with retroviral vectors that gave insertional mutagenesis at the p53 locus. We presume this line acquired addition errors at crisis.

184AA5-7

These are immortal lines that arose in finite lifespan BaP post-stasis 184Aa populations transduced with control retroviruses. They have not been characterized, but their early rapid growth suggests they are p53(-) like 184AA2 and 184AA3, and are a consequence of insertional mutagenesis. These cells are not routinely distributed.

184AA8

This is a new line of indefinite lifespan that emerged from BaP post-stasis 184Aa. It is not yet well-characterized; it’s gradual conversion process is consistent with a p53(+) status. It is not yet available for distribution.

184AaGS1-2

Finite lifespan BaP post-stasis 184Aa was transduced with GSE22 and produced two immortally transformed cultures. Immortalization is clonal, and each isolate is likely to be somewhat different. We presume these lines have acquired addition errors at crisis. These lines have not been well-characterized and are not routinely distributed.

184AaGS1 has AIG.

184AaMY1-5 (Garbe et al. 2014)
Finite lifespan BaP post-stasis 184Aa was transduced with c-myc on multiple occasions; each time there has been efficient rapid immortalization of the population. These lines have not been well-characterized. They are not routinely distributed but can be made available.

Finite lifespan BaP post-stasis 184Aa was transduced with the breast cancer-associated oncogene ZNF217 on multiple occasions. Immortalization is clonal, and each isolate is likely to be somewhat different. We presume these lines have acquired additional errors during the period of agonescence. The lines examined are p53(+). They are not routinely distributed but can be made available.

Unlimited
quantities of later passages of this immortal line are available for
distribution. More limited quantities of
early passage newly immortal (pre- and mid- conversion) cells are available
upon specific request. Clonal isolates
are also available. These cells are wild type for p53, and RB, and are not
anchorage independent or tumorigenic. They have a low level of karyotypic instability. We previously thought that this line was derived from BaP post-stasis 184Be, however recent studies (Severson et al. and in prep) indicate that it arose from a minor (~10%) population in the 184Be culture, which we have now named 184Bd. Unfortunately, pure cultures of 184Bd are not available.

This is a new line of indefinite lifespan that emerged from BaP post-stasis 184Be. It is not yet well-characterized; it’s gradual conversion process is consistent with a p53(+) status. This line is not routinely distributed but can be made available.

184BE2This is a new line of indefinite lifespan that emerged from BaP post-stasis 184Be. It is not yet well-characterized; it’s gradual conversion process is consistent with a p53(+) status. It is not yet available for distribution.

Finite lifespan BaP post-stasis 184Be was transduced with c-myc, producing efficient rapid immortalization of the population. This line has not been well-characterized and is not yet available for distribution.

Finite lifespan BaP post-stasis 184Ce was transduced with c-myc, producing efficient rapid immortalization of the population. This line has not been well-characterized and is not yet available for distribution. We have not derived any “spontaneous” lines from 184Ce (culture of 184Ce has been more limited than 184Aa or 184Be).

Pre-stasis 184F grown in MM
were transduced with hTERT at passage 3. A clonal population maintained growth
after the
vast majority of the cells ceased growth at stasis.The population
gradually lost expression of p16 while gaining resistance to TGFß growth
inhibition.These cells have not been
well-characterized. This line is not routinely distributed but can be
made available.

Finite lifespan
post-selection 184 was transduced with c-myc on multiple occasions.In 10 independent experiments 1 clonal
immortal line emerged.We presume this
line acquired additional errors during the period of agonescence.These cells have not been well-characterized.
This line is not routinely distributed but can be made available.

Finite
lifespan post-selection 184 was transduced with ZNF217 on multiple
occasions.In 7 independent experiments 4 clonal immortal lines
emerged;
each isolate is likely to be somewhat different.We presume these lines acquired additional
errors during the period of agonescence. The lines examined are p53(+).These lines are not routinely distributed but can be
made available.

Finite
lifespan post-selection 184 was transduced with ZNF217, followed by c-myc, on
multiple occasions. In 5 independent experiments 4 clonal immortal lines
emerged.Three of these lines emerged
soon after the exposure to c-myc, prior to agonescence; Southern analysis
indicated they are clonal. The 2 lines
examined by array CGH show no alterations in copy numbers.We presume no additional genomic mutations
were required for immortalization. These
lines are
not routinely distributed but can be made available on a collaborative basis
upon request.

184ZNMY3-N

We
have transduced 184ZNMY2 and 184ZNMY3 with oncogenic erbB2 (neu).These lines then acquired AIG. CGH analysis
shows no copy number alterations.

184ZNGS1Finite lifespan post-selection 184 was transduced with ZNF217, followed byGSE22, in two experiments. In 4 independent exposures one immortal line emerged soon after the exposure to GSE22, prior to agonescence. These cells have not been well-characterized. This line is not routinely distributed but can be made available.

Transduction
of hTERT into post-selection p16(-) HMEC producesefficientimmortalization bypassing the genomic instability of agonescence,
andconversion. Unlimited quantities of 184BTERT are available for
distribution.

2C) Line derived from post-selection HMEC transduced with HPV-E6

184-E6

(Garbe et al. 1999)Transduction of the HPV16 E6 gene into post-selection p16(-) HMEC produces common immortalization following crisis, bypassing conversion. These cells are not routinely distributed but can be made available.